Protein folding, stability and function are driven by noncovalent interactions. Consequently, native proteins at physiological temperatures continuously sample many conformations or conformational substrates. The populations of these conformational substrates and their kinetics of interconversion are manifested directly in protein folding, stability, ligand binding, catalysis, gating of ion channels, and allosteric effects. Prion diseases and other diseases of protein deposition are some of the most compelling evidence for protein conformational dynamics and their impact on human health. In these diseases, proteins are driven from their native conformations to non-native conformations and aggregation by mutation or the presence of misfolded protein. Conformational dynamics are also of considerable interest and concern in the area of drug design. The proposed research will use of an array of experimental and computational tools to develop a quantitative understanding of how conformation, stability, and dynamics are related in ubiquitin. A major focus is amide hydrogen (NH) exchange, which potentially provides information regarding the thermodynamics and kinetics of protein motions at individual residues in native proteins. The first two aims will establish a more precise understanding of the molecular basis for slow NJ exchange in native proteins. Aims 3 and 4 will expand investigation into molecular motions in native ubiquitin using both NH exchange and NMR relaxation studies. Prospects for success on all four aims are enhanced by collaborations with experts in molecular dynamics, mass spectrometry, stopped-flow experiments, and x-ray crystallography.